Trap Mass Spectrometer Research Articles

Simultaneous detection of sibutramine and phenolphthalein in a diet jelly health food product that caused health problems.

The Public Health Center in Chiba Prefecture, Japan, received a consultation from a resident of Chiba Prefecture who consumed a diet jelly health food product and experienced health problems. To investigate the cause of the health problems, we examined the twofood products for the presence of pharmaceutical ingredients. A screening analysis using ultra-high-performance liquid chromatography with a photodiode array detector (UPLC-PDA) indicated the presence of sibutramine and phenolphthalein in the food product. Analysis using an ultra-high-performance liquid chromatography-quadrupole-Kingdon trap mass spectrometer (UHPLC-Q-Kingdon trap MS) confirmed the presence of sibutramine and phenolphthalein. Quantitative analysis using UPLC-PDA showed that sibutramine and phenolphthalein were present at 15 and 16mg/bag and 2.4 and 2.6mg/bag, respectively. According to the drug insert for sibutramine capsules in the United States, the recommended medicinal dose of sibutramine should not exceed 15mg/day, and the amount ingested in the present case exceeded that value. The present study results indicated that ingestion of the jelly health food product may cause health problems.

Development and validation of an analytical method for the determination of direct oral anticoagulants (DOAC) and the direct thrombin-inhibitor argatroban by HPLC–MS/MS

Since direct oral anticoagulants (DOAC) are administered frequently to an elderly, co-morbid population, medical emergencies including trauma, acute bleeding or organ failure are not uncommon. In these situations, the type, dosage or the time of last intake of anticoagulants is often unknown and single substance analysis by functional tests is only possible if the substance contained in the sample is known. A reliable and validated toxicology screen of DOAC and argatroban would be helpful inform not only attending physicians in the emergency department but also law enforcement and courts of justice. After precipitation with acetone, HPLC separation was achieved on a Phenomenex Luna Pentafluorophenyl Colum using acetonitrile–water (90:10, v/v) as mobile phase system. Detection was performed using a 3200 Q Trap mass spectrometer (AB Sciex). For analysis MRM Scans (MS/MS) with positive ionization were chosen. The method was validated for blank serum as the matrix of choice. Limits of detection are between 0.5 and 1.0 ng/mL, limits of quantification are between 1.9 and 3.6 ng/mL and recoveries are above 60%. The applicability of the method was demonstrated by the determination of DOAC in body fluids from forensic cases and in therapeutic drug monitoring. The rapid simultaneous detection and quantification of apixaban, argatroban, dabigatran etexilate, dabigatran, edoxaban and rivaroxaban in body fluids by HPLC–MS/MS closes an important gap in emergency toxicology.

Bioassay-Guided Interpretation of Antimicrobial Compounds in Kumu, a TCM Preparation From Picrasma quassioides' Stem via UHPLC-Orbitrap-Ion Trap Mass Spectrometry Combined With Fragmentation and Retention Time Calculation.

The stem of Picrasma quassioides (PQ) was recorded as a prominent traditional Chinese medicine, Kumu, which was effective for microbial infection, inflammation, fever, and dysentery, etc. At present, Kumu is widely used in China to develop different medicines, even as injection (Kumu zhusheye), for combating infections. However, the chemical basis of its antimicrobial activity has still not been elucidated. To examine the active chemicals, its stem was extracted to perform bioassay-guided purification against Staphylococcus aureus and Escherichia coli. In this study, two types of columns (normal and reverse-phase) were used for speedy bioassay-guided isolation from Kumu, and the active peaks were collected and identified via an UHPLC-Orbitrap-Ion Trap Mass Spectrometer, combined with MS Fragmenter and ChromGenius. For identification, the COCONUT Database (largest database of natural products) and a manually built PQ database were used, in combination with prediction and calculation of mass fragmentation and retention time to better infer their structures, especially for isomers. Moreover, three standards were analyzed under different conditions for developing and validating the MS method. A total of 25 active compounds were identified, including 24 alkaloids and 1 triterpenoid against S. aureus, whereas only β-carboline-1-carboxylic acid and picrasidine S were active against E. coli. Here, the good antimicrobial activity of 18 chemicals was reported for the first time. Furthermore, the spectrum of three abundant β-carbolines was assessed via their IC50 and MBC against various human pathogens. All of them exhibited strong antimicrobial activities with good potential to be developed as antibiotics. This study clearly showed the antimicrobial chemical basis of Kumu, and the results demonstrated that HRMS coupled with MS Fragmenter and ChromGenius was a powerful tool for compound analysis, which can be used for other complex samples. Beta-carbolines reported here are important lead compounds in antibiotic discovery.

Simultaneous determination of selected catechins and pyrogallol in deer intoxications by HPLC-MS/MS

A selective and sensitive analytical method for the determination of selected catechins (catechin, epicatechin, gallocatechin, epigallocatechin) and pyrogallol in biological matrices by HPLC-MS/MS was developed. The utilized sample preparation technique was a two-stage liquid–liquid extraction using ethyl acetate. The HPLC-system was equipped with a Phenomenex Luna Pentafluorophenyl Column (150 × 2 mm, 5 µm) and operated with an acetonitrile–water gradient as a mobile phase system. Detection was performed with a 3200 Q Trap mass spectrometer. For analysis the mass spectrometer was used in the MRM-mode with negative ionization.The method validation was performed with serum as matrix. The selectivity of the method as well as the linearity of calibration was successfully proven for all analytes. The limits of quantification were between 5.3 and 11.2 ng/mL and the recovery rates were above 50 % for all analytes.Results from the samples of three deer poisoning cases demonstrated that the developed HPLC-MS/MS method is applicable to real biological samples.

Increased Single-Spectrum Top-Down Protein Sequence Coverage in Trapping Mass Spectrometers with Chimeric Ion Loading.

Fourier transform mass spectrometers routinely provide high mass resolution, mass measurement accuracy, and mass spectral dynamic range. In this work, we utilize 21 T Fourier transform ion cyclotron resonance (FT-ICR) to analyze product ions derived from the application of multiple dissociation techniques and/or multiple precursor ions within a single transient acquisition. This ion loading technique, which we call, "chimeric ion loading", saves valuable acquisition time, decreases sample consumption, and improves top-down protein sequence coverage. In the analysis of MCF7 cell lysate, we show collision-induced dissociation (CID) and electron-transfer dissociation (ETD) on each precursor on a liquid chromatography-mass spectrometry (LC-MS) timescale and improve mean sequence coverage dramatically (CID-only 15% vs chimeric 33%), even during discovery-based acquisition. This approach can also be utilized to multiplex the acquisition of product ion spectra of multiple charge states from a single protein precursor or multiple ETD/proton-transfer reactions (PTR) reaction periods. The analytical utility of chimeric ion loading is demonstrated for top-down proteomics, but it is also likely to be impactful for tandem mass spectrometry applications in other areas.

VGAM: Compact and Low‐Power Mass Spectrometer‐Based Instrumentation for Volcanic Gas Monitoring

AbstractCompact mass spectrometers (MSs) can provide simultaneous multispecies analysis with high sensitivity and precision. For volcanic gas monitoring in situ, instrumental mass spectrometry requires reliability and ruggedness combined with low power usage and portability. The Volcanic Gas Analytical Monitor (VGAM) is capable of quantitative molecular analysis of a variety of atmospheric and volcanic gases in a single sensor by ion trap mass spectrometry. These gases include H2, He, H2O, N2, O2, Ar, NO, N2O, CO, CO2, H2S, SO, SO2, and CH4. Unlike previous field instruments using magnetic sector and quadrupole MSs with vacuums backed by compact turbomolecular pumps, the VGAM uses a low‐power autoresonant ion trap MS and nonevaporable getter‐ion pumped vacuum that operates at only 25‐W total power, which is often the power requirement for just the MS. Ratiometric mass spectral response is combined with total pressure measurements to report absolute partial pressures. Data are generated in real time and are recorded to internal flash memory. Relatively low power (<1 W) data telemetry to remote sites is possible. Analysis of volcanic plumes, fumaroles, and solfatara fields is accomplished by direct inlet of gases, after trapping as much excess water vapor in both the external and internal foreline as possible. We report herein on the VGAM autorun and postprocessing procedures, initial calibration results for CO2, and the results of a brief field deployment at Sulfur Banks solfatara field, Kilauea Volcano, Hawaii.

Response of QIT-MS to Noble Gas Isotopic Ratios in a Simulated Venus Flyby

The primary objective of the present study is to investigate the science return of future Venus atmosphere probe mission concepts using the Quadrupole Ion Trap (QIT) Mass Spectrometer (MS) Instrument (QIT-MS-I). We demonstrate the use of Monte-Carlo simulations in determining the optimal ion trapping conditions and focus the analysis on retrieving isotope ratios of noble gases in the model sample of the Venus atmosphere. Sampling takes place at a constant velocity of ~10 km/s between 112–110 km altitude and involves the use of getter pumps to remove all chemically-active species, retaining inert noble gases. The enriched sample is leaked into passively pumped vacuum chamber where it is analyzed by the QIT-MS sensor (QIT-MS-S) for 40 minutes. The simulated mass spectrum, as recorded by the QIT-MS-S, is deconvoluted using random walk algorithm to reveal relative abundances of noble gas isotopes. The required precision and accuracy of the deconvolution method is benchmarked against the a priori known model composition of the atmospheric sample.

Biomolecular Clusters Distribution up to Mega Dalton Region Using MALDI-Quadrupole Ion Trap Mass Spectrometer.

We present the first report on complete cluster distributions of cytochrome c (molecular weight of 12.4 kDa) and bovine serum albumin ((BSA), molecular weight of 66.4 kDa) with mass-to-charge ratio (m/z) reaching 350,000 and 1,400,000, respectively, by matrix-assisted laser desorption/ionization (MALDI). Large cluster distributions of the analytes were measured by our homemade frequency-scanned quadrupole ion trap (QIT) mass spectrometer with a charge detector. To our knowledge, we report the highest m/z clusters of these two biomolecules. The quantitative results indicate that large clusters ions of cytochrome c and BSA follow the power law (r2 > 0.99) with cluster size distribution, which provides experimental evidence for the laser ablation studies of MALDI.

Online eluent-switching technique coupled anion-exchange liquid chromatography–ion trap tandem mass spectrometry for analysis of non-steroidal anti-inflammatory drugs in pig serum

A novel method for online extraction, pH-gradient separation, and analysis of nine non-steroidal anti-inflammatory drugs (NSAIDs) was developed by coupling online eluent-switching technique to single anion-exchange chromatographic column/ion trap mass spectrometer (MS) and used for monitoring NSAIDs residues in pig serum. A neutral eluent and a pH-gradient eluent were used for extraction and separation of NSAIDs, respectively. Each of nine NSAIDs has an MS precursor ion of either [M−H]− or [M−Na]−. The extracted ion chromatogram for a specific product ion of each NSAID was used for its quantitative analysis. The dynamic linear ranges of calibration curves were all 0–200ngmL−1 (R2>0.9950). The analysis accuracies estimated by spiking standard concentrations at 20, 100, and 200ngmL−1 were 80.5–99.9%. The corresponding intra-day and inter-day precisions (RSD%) were 2.5–14.5% and 2.9–15.2%, respectively. The limit of detection/limit of quantitation of NSAIDs were 1.3/4.3, 0.5/1.6, 0.2/0.5, 2.5/8.2, 1.5/4.9, 0.6/2.1, 0.6/2.0, 0.5/1.7, and 0.6/2.1ngmL−1 for carprofen, diclofenac, flunixin, ibuprofen, ketoprofen, meclofenamic acid sodium, mefenamic acid, niflumic acid, and tolfenamic acid, respectively. After 1h injection of a dose containing 2mgkg−1 weight pig of flunixin and tolfenamic acid to the pigs, a residue amount of 3480±36ngmL−1 and 431±13ngmL−1, respectively, was reached for the incurred pig serum specimens and both residues were reduced to about 20ngmL−1 at the time of 24h.

Development and validation of an universal interface for compound-specific stable isotope analysis of chlorine (37Cl/35Cl) by GC-high-temperature conversion (HTC)-MS/IRMS.

A universal application of compound-specific isotope analysis of chlorine was thus far limited by the availability of suitable analysis techniques. In this study, gas chromatography in combination with a high-temperature conversion interface (GC-HTC), converting organic chlorine in the presence of H2 to gaseous HCl, was coupled to a dual-detection system, combining an ion trap mass spectrometer (MS) and isotope-ratio mass spectrometer (IRMS). The combination of the MS/IRMS detection enabled a detailed characterization, optimization, and online monitoring of the high-temperature conversion process via ion trap MS as well as a simultaneous chlorine isotope analysis by the IRMS. Using GC-HTC-MS/IRMS, chlorine isotope analysis at optimized conversion conditions resulted in very accurate isotope values (δ(37)Cl(SMOC)) for measured reference material with known isotope composition, including chlorinated ethylene, chloromethane, hexachlorocyclohexane, and trichloroacetic acids methyl ester. Respective detection limits were determined to be <15 nmol Cl on column with achieved precision of <0.3‰.

Mass Spectrometry-based Workflow for Accurate Quantification of Escherichia coli Enzymes: How Proteomics Can Play a Key Role in Metabolic Engineering

Metabolic engineering aims to design high performance microbial strains producing compounds of interest. This requires systems-level understanding; genome-scale models have therefore been developed to predict metabolic fluxes. However, multi-omics data including genomics, transcriptomics, fluxomics, and proteomics may be required to model the metabolism of potential cell factories. Recent technological advances to quantitative proteomics have made mass spectrometry-based quantitative assays an interesting alternative to more traditional immuno-affinity based approaches. This has improved specificity and multiplexing capabilities. In this study, we developed a quantification workflow to analyze enzymes involved in central metabolism in Escherichia coli (E. coli). This workflow combined full-length isotopically labeled standards with selected reaction monitoring analysis. First, full-length (15)N labeled standards were produced and calibrated to ensure accurate measurements. Liquid chromatography conditions were then optimized for reproducibility and multiplexing capabilities over a single 30-min liquid chromatography-MS analysis. This workflow was used to accurately quantify 22 enzymes involved in E. coli central metabolism in a wild-type reference strain and two derived strains, optimized for higher NADPH production. In combination with measurements of metabolic fluxes, proteomics data can be used to assess different levels of regulation, in particular enzyme abundance and catalytic rate. This provides information that can be used to design specific strains used in biotechnology. In addition, accurate measurement of absolute enzyme concentrations is key to the development of predictive kinetic models in the context of metabolic engineering.

Structural characterization of cyclosporin A, C and microbial bio-transformed cyclosporin A analog AM6 using HPLC-ESI-ion trap-mass spectrometry.

Cyclosporin A (CyA), a cyclic undecapeptide produced by a number of fungi, contains 11 unusual amino acids, and has been one of the most commonly prescribed immunosuppressive drugs. To date, there are over sixty different analogs reported as congeners and analogs resulting from precursor-directed biosynthesis, human CYP-mediated metabolites, or microbial bio-transformed analogs. However, there is still a need for more structurally diverse CyA analogs in order to discover new biological potentials and/or improve the physicochemical properties of the existing cyclosporins. As a result of the complexity of the resulting mass spectrometric (MS) data caused by its unusual amino acid composition and its cyclic nature, structural characterization of these cyclic peptides based on fragmentation patterns using multiple tandem MS analyses is challenging task. Here, we describe, an efficient HPLC-ESI-ion trap MS(n) (up to MS(8)) was developed for the identification of CyA and CyC, a (Thr(2))CyA congener in which L-aminobutyric acid (Abu) is replaced by L-threonine (Thr). In addition, we examined the fragmentation patterns of a CyA analog obtained from the cultivation of a recombinant Streptomyces venezuelae strain fed with CyA, assigning this analog as (γ-hydroxy-MeLeu(6))CyA (otherwise, known as an human CYP metabolite AM6). This is the first report on both the MS(n)-aided identification of CyC and the structural characterization of a CyA analog by employing HPLC-ESI-ion trap MS(n) analysis.

An Interplay Between Infrared Multiphoton Dissociation Fourier-Transform Ion Cyclotron Resonance Mass Spectrometry and Density Functional Theory Computations in the Characterization of a Tripodal Quinolin-8-Olate Gd(III) Complex

A new hexadentate, tripodal 8-hydroxyquinoline based ligand (QH3) and its gadolinium(III) tris-chelated (GdQ) complex with hemicage structure was investigated by using high resolution Fourier-transform ion cyclotron resonance mass spectrometry (FTICRMS). The protonated adduct of the free ligand and its hemicage tripodal Gd(III) complex, [GdQ + H](+), were first observed in experiments of electrospray ionization (ESI) with a linear ion trap (LTQ) mass spectrometer and further investigated by using high resolution FTICRMS. Gas-phase dissociation of the protonated Gd(III) complex, by infrared multiphoton dissociation (IRMPD) FTICR MS, demonstrated a fragmentation pattern with six main product cluster ions labeled as [Fn](+) (n = 1 up to 6). These product ions suggest the elimination of 7-amino-alkyl or 7-alkyl chains of the hemicage moiety. High resolution MS conditions allowed the elucidation of the fragmentation pattern and product ion structures along with the determination, among the isotopic pattern of Gd, of the chemical compositions of closely related species, which differ in terms of hydrogen content. Among the Gd six naturally stable isotopes, (158)Gd is the most abundant, and its peak within each cluster was used as a reference for distinguishing each product ions. Computational DFT investigations were applied to give support to some hypothesis of fragmentation pathways, which could not have been easily justified on the basis of the experimental work. Furthermore, computational studies suggested the coordination geometry of the protonated parent complex and the five- and four-coordinated complexes, which derive from its fragmentation. Furthermore, experimental and computational evidences were collected about the octet spin state of the parent compound.

Determination of nine β-agonist residues in pig tissues by liquid chromatography-tandem mass spectrometry combining with library search

A new method has been developed using a hybrid triple-quadrupole linear ion trap (QTrap) mass spectrometer for the fast detection and identification of nine beta-agonists, clenbuterol, salbutamol, ractopamine, ritodrine, terbutaline, isoxsuprine, tulobuterol, cimaterol and bambuterol, in one single liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. The homogenized tissue samples were purified with liquid-liquid extraction after enzymatic hydrolysis by P-glucuronidase/aryl sulfatase. After gradient elution separation on C(18) LC column using acetonitrile and formic acid aqueous solution as the mobile phases, a multiple reaction monitoring (MRM) scan as survey scan and an enhanced product ion (EPI) scan as dependent scan were performed in an information dependent acquisition (IDA) experiment. Finally, the identification of the drugs was carried out by library search with a newly developed MS/MS library based on EPI spectra at three different collision energies in positive mode. The analytical method in the present study was well validated and good results were obtained with respect to precision, repeatability and spiked recovery. The limits of detection of residues were 0.1 -0.2 micro g/kg for beta-agonists, and with a linear range from 0.1 to 50.0 micro g/L. Three concentration levels of 0. 5, 1. 0 and 5. 0 pg/kg were spiked in pig tissues, and the overall recoveries were between 72.0% and 95.1% with the relative standard deviations (RSDs) between 3. 1% and 12.1%. The real sample test showed that this method could be used for sensitive and accurate determination of beta-agonist residues in pig tissue

The aroma biogenesis-related Olea europaea ALCOHOL DEHYDROGENASE gene is developmentally regulated in the fruits of two O. europaea L. cultivars

In plants, the aroma imprint is expressed through well-established and constant ratios between different compounds, which mainly develop through the Lipoxygenase pathway. Such imprint strongly relies on plant genetic traits but can be modulated by exogenous environmental factors. In order to enlarge knowledge on the genetic control underlying aroma biogenesis in olive (Olea europaea L.) drupes, in the present study we characterized one member of ALCOHOL DEHYDROGENASE gene family. The ALCOHOL DEHYDROGENASE activities act along Lipoxygenase enzymatic cascade by interconverting aldehydes into alcohols. The temporal expression pattern of O. europaea ALCOHOL DEHYDROGENASE was monitored by quantitative Real Time-Polymerase Chain Reaction in drupes at different developmental stages (i.e. from light green phase to ripening stage) of two olive cultivars widely used for olive and oil production. The expression of a known LIPOXYGENASE gene (ACG56281.1) was jointly monitored as marker of drupe development. Results were analysed in relation to the spectrum of volatile compounds developed from crushed drupes estimated by Solid-Phase Micro-Extraction method in-line with a Gas Chromatograph/ion trap Mass Spectrometer. The obtained results showed that both cultivar and maturation stage impact on the expression of O. europaea ALCOHOL DEHYDROGENASE gene which in the pathway is co-ordinately modulated with LIPOXYGENASE expression. The transcriptional differences between cultivars appeared to be related to variation of metabolite contents at early developmental stages. The joint role of genes acting in the Lipoxygenase pathway as molecular markers for flavour characteristic of olive drupes is discussed.

Abstract 4795: Metabolic biomarkers of hepatocellular carcinoma

Abstract Characterizing the metabolic changes pertaining to hepatocellular carcinoma (HCC) in patients with liver cirrhosis (LC) is believed to contribute towards early detection, treatment, and our understanding of the molecular mechanisms of HCC. Prospectively evaluated adult patients with LC were recruited from our hepatology practice. All patients were diagnosed to have LC on the basis of established clinical, laboratory and imaging criteria. Cases were diagnosed to have HCC based on either histology or characteristic imaging features. The LC controls were required to be HCC free for at least 6 months from the time of study entry. We compared metabolite levels in sera of 78 HCC cases with 184 LC controls by using ultra performance liquid chromatography coupled with a hybrid quadrupole time-of-flight mass spectrometry (UPLC-QTOF MS). Following data preprocessing, the most relevant ions in distinguishing HCC cases from LC controls were selected by statistical methods. Putative metabolite identifications for these ions were obtained through mass-based database search. Pathway analysis of these putative identifications reveals significant enrichment of metabolites involved in bile acid biosynthesis, porphyrin and chlorophyll metabolism, and arachidonic acid metabolism. Verification of the identities of selected metabolites was conducted by comparing their MS/MS fragmentation patterns and retention time with those from authentic compounds. Quantitation of these metabolites was performed in a subset of the serum samples (10 HCC and 10 LC) using isotope dilution by selected reaction monitoring (SRM) on triple quadrupole linear ion trap (QqQLIT) mass spectrometer. The results of this analysis confirms that metabolites involved in phospholipid catabolism such as sphingosine-1-phosphate (S-1-P) and lysophosphatidylcholine are up-regulated in sera of HCC vs. those with LC. Down-regulated metabolites include those involved in cholesterol metabolism such as glycochenodeoxycholic acid 3-sulfate (3-sulfo-GCDCA), glycocholic acid (GCA), glycodeoxycholic acid (GDCA), taurocholic acid (TCA), taurochenodeoxycholate (TCDCA), as well as those involved in heme catabolism including billuribin, I-Urobili, and bilirubin glucuronide. The preliminary results obtained in this study provide useful insights into our understanding of metabolic differences between HCC and LC. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4795. doi:1538-7445.AM2012-4795

Gas-phase fluorescence excitation and emission spectroscopy of mass-selected trapped molecular ions

A flexible interface to perform optical spectroscopic measurements on gaseous ions stored in a modified commercial quadrupole ion trap (QIT) mass spectrometer is described. The modifications made to the mass spectrometer did not adversely affect its operating characteristics. Gas-phase ions are produced using electrospray ionization, mass isolated and stored in the trapping mass spectrometer. The ions are subsequently irradiated with visible light from a tunable laser and dispersed fluorescence spectra are recorded simultaneously. Mass spectra are recorded after the irradiation period. This set-up allows us to track a range of possible outcomes upon photoexcitation of selected ions including fluorescence, photofragmentation and photodetachment of electrons. The experimental set-up is characterized using rhodamine 590, which is a methyl ester variant of rhodamine 6G. Fluorescence excitation and emission spectra of gaseous rhodamine 590 are measured and compared with solution-phase spectra. Excitation and emission maxima for the gaseous ions are found to lie at higher energy than for the solvated rhodamine 590. In addition, the gas-phase Stokes shift is significantly smaller than the solution-phase Stokes shift. The effects of several experimental parameters on the observed fluorescence signal are investigated, including laser power, relative number of ions, q(z) trapping parameter and buffer gas pressure. In addition to its use for the photophysical characterization of the intrinsic properties of ionic chromophores, this set-up may be used to investigate the properties of mass-selected, dye-labeled biomolecules, both alone and in well-defined complexes and clusters.

Serum Levels of Risperidone and Its Metabolite, 9-Hydroxyrisperidone: Correlation Between Drug Concentration and Clinical Response

The aim of this study was to assess a method able to analyze serum levels of risperidone (RIS) and its metabolite, 9-hydroxyrisperidone (9-OH-RIS), and to investigate possible relationships between changes in serum concentrations of these drugs and clinical measures, so to identify early markers of treatment response. The authors developed a sensitive and specific liquid chromatography-tandem mass spectrometry method to measure RIS and its metabolite in serum. Fifteen RIS-naive patients were admitted to an acute psychiatric care unit and treated with 4-6 mg/d oral RIS. At days 7 and 21 of hospital stay, serum levels were measured; clinical scales and serum prolactin were assessed. RIS and its metabolite were analyzed by a Q-Trap 2000 triple quadrupole/ion trap mass spectrometer in the multiple reaction-monitoring mode. Chromatographic separation was accomplished using a cyano column with an analytical run of 9 minutes. The calibration curve exhibited consistent linearity and reproducibility in the range 0-100 ng/mL for both analytes. Lower limit of quantification was 0.2 ng/mL; limit of detection, for a signal to noise ratio of 3, was 0.05 ng/mL for both analytes. Serum RIS and 9-OH-RIS levels increased at day 7, reaching a steady state, and remaining constant up to day 21. Scores on psychopathology rating scales decreased; serum prolactin and neurological rating scale for extrapyramidal effects rose at day 7 and remained stable thereafter. No correlation was found between serum concentration values, including sum and ratio of RIS and 9-OH-RIS, and any of the other clinical values (serum prolactin and clinical scales). These data indicate that clinical changes are related to the achievement of steady state levels of RIS and its metabolite and are maintained, but not continued, with continued RIS treatment. Therapeutic drug monitoring of RIS and its metabolites is not recommended as a routine procedure in patients with psychotic disorders.

A Liquid Chromatography-Tandem Mass Spectrometry Method for the Determination of 5-Fluorouracil Degradation Rate by Intact Peripheral Blood Mononuclear Cells

5-Fluorouracil (5-FU) is a major chemotherapy drug used for the treatment of tumors. It is catabolized mainly by dihydropyrimidine dehydrogenase, and patients with a complete or partial deficiency of dihydropyrimidine dehydrogenase activity are at risk of developing severe 5-FU-associated toxicity. The aim of this study was to demonstrate that intact peripheral blood mononuclear cells (PBMCs) can be an effective model to evaluate the degradation rate of 5-FU. We developed a sensitive and specific liquid chromatography-tandem mass spectrometry method to measure in vitro the rate of 5-FU degradation by intact PBMC. 5-FU degradation rate was determined by measuring the decrease of a fixed amount of 5-FU (10 microg/mL) added to a solution of PBMC, after 2 hours incubation, expressed as nanogram per milliliter of 5-FU degraded per minute x 10(6) cells. Freshly prepared intact PBMC can degrade efficiently in vitro-added 5-FU. The assay consists of 3 steps: (1) PBMC isolation from peripheral blood, (2) PBMC incubation with 5-FU in vitro for different times, and (3) determination of 5-FU amount to calculate the degradation rate. 5-FU was analyzed by a Q Trap 2000 triple quadrupole/ion trap mass spectrometer in the multiple-reaction-monitoring modes. The chromatographic separation was accomplished using a C18 column with a run time of 16 minutes. By analyzing samples from 39 patients with no 5-FU toxicity, the mean 5-FU degradation rate was 1.85 +/- 0.50 ng x mL(-1) x min(-1) x 10(6) cells. The assessment of a test to measure 5-FU degradation rate in PBMC of patients before 5-FU administration could represent a prescreening method for evaluating the possible toxicity of this drug as an aid to set up a personalized medicine approach for each patient.

Vibrational spectroscopy of bare and solvated ionic complexes of biological relevance

The low density of ions in mass spectrometers generally precludes direct infrared (IR) absorption measurements. The IR spectrum of an ion can nonetheless be obtained by inducing photodissociation of the ion using a high-intensity tunable laser. The emergence of free electron lasers (FELs) and recent breakthroughs in bench-top lasers based on nonlinear optics have now made it possible to routinely record IR spectra of gas-phase ions. As the energy of one IR photon is insufficient to cause dissociation of molecules and strongly bound complexes, two main experimental strategies have been developed to effect photodissociation. In infrared multiple-photon dissociation (IR-MPD) many photons are absorbed resonantly and their energy is stored in the bath of vibrational modes, leading to dissociation. In the "messenger" technique a weakly bound van der Waals atom is detached upon absorption of a single photon. Fundamental, historical, and practical aspects of these methods will be presented. Both of these approaches make use of very different methods of ion preparation and manipulation. While in IR-MPD ions are irradiated in trapping mass spectrometers, the "messenger" technique is generally carried out in molecular beam instruments. The main focus of this review is the application of IR spectroscopy to biologically relevant molecular systems (amino acids, peptides, proteins). Particular issues that will be addressed here include gas-phase zwitterions, the (chemical) structures of peptides and their collision-induced dissociation (CID) products, IR spectra of gas-phase proteins, and the chelation of metal-ligand complexes. Another growing area of research is IR spectroscopy on solvated clusters, which offer a bridge between the gas-phase and solution environments. The development of state-of-the-art computational approaches has gone hand-in-hand with advances in experimental techniques. The main advantage of gas-phase cluster research, as opposed to condensed-phase experiments, is that the systems of interest can be understood in detail and structural effects can be studied in isolation. It will be shown that IR spectroscopy of mass-selected (bio)molecular systems is now well-placed to address specific questions on the individual effect of charge carriers (protons and metal ions), as well as solvent molecules on the overall structure.